Vertebrate opsins in photoreceptors and the retinal pigment epithelium (RPE) have fundamental roles in the visual process. The visual pigments in photoreceptors are bound to 11 -cis-retinal and are responsible for the initiation of visual excitation. Retinochrome-like opsins in the RPE are bound to all-trans-retinal and may play an important role in chromophore metabolism. The retinal G protein-coupled receptor (RGR) in the RPE and Muller cells is an opsin that is necessary for normal synthesis of 11 -cis-retinal and regeneration of rhodopsin during light adaptation. It is hypothesized that RGR has a central role in light-dependent synthesis of 11-cis-retinal and the regeneration of rhodopsin, hence RGR is involved in the visual cycle. The objective of this grant application is to prove this hypothesis by demonstration of a detailed mechanistic model for the function of RGR at the molecular and physiological levels. The study of RGR opsin is impeded by limitations in accessibility, low abundance, biochemical properties, and the lack of an appropriate cell culture model. To be able to study the RGR opsin, three experimental approaches to this research problem have been created. A functional RGR opsin from bovine RPE has been isolated. Second, a cell culture model of RPE cells that stably express RGR and remain able to process retinoids has been established. Third, the RGR opsin knockout mouse with an informative phenotype has been produced. These experimental approaches will be used in this grant proposal to further investigate the function of RGR and its role in retinoid metabolism and regulation in the RPE. By preserving rhodopsin levels and retinal sensitivity, the RGR opsin gene helps to prevent night blindness and provides a selective advantage for species subject to wide variation in environmental luminance. Further understanding of RGR at the biochemical level will be important in learning how RPE cells work, and characterization of defects in RGR function may show how human RPE cells undergo dysfunction and deteriorate leading to disease. Since RGR is involved in a fundamental visual process, defects in RGR are likely to impair the health of the RPE and retina. The importance of RGR opsin to the health and viability of the neuroretina is shown by mutations in the human RGR gene that segregate with retinitis pigmentosa (RP) in patients with autosomal dominant or recessive RP. Errors in retinoid metabolism in either the photoreceptors or RPE cells may lead to abnormal levels of A2E, a major compound of lipofuscin that accumulates in the RPE during aging and age-related macular degeneration.